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Nitza Goldenberg-Cohen, Tamilla Sadikov, Bat Chen R. Avraham-Lubin, Nadir Askenasy; Intravitreal Injection of Adult Bone Marrow Derived Stem Cells To Developing Retina of Newborn Mice. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3177.
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Bone marrow cells have been shown to home to sites of injury and participate in the process of remodeling after injury. The smallest population of nucleated bone marrow cells has been shown to differentiae into a range of epithelial tissues, liver, endothelium and insulin-producing cells, in addition to long-term hematopoietic reconstituting potential. The retina, under physiological conditions, continues to develop 3 weeks post partum, to complete neurogenesis and angiogenesis. To determine whether these cells can contribute to the retina in the absence of injury and inflammation, we conceived that the neonatal eye would be the most favorable environment for such incorporation and differentiation. Therefore, we seek to determine the participation of this subset of candidate multipotent cells to retinal modeling after birth.
Adult bone marrow cells were isolated by counterflow centrifugal elutriation that separates a subset accounting for 8-14% of the bone marrow cells at a flow rate of 25 ml/min (Fr25). To obtain a hematopoietic lineage-negative fraction (Fr25lin-), immunomagnetic depletion was performed for Ter-119 (erythrocytes), GR-1 (granulocytes), Mac-1 (macrophages), NK1.1 (natural killer cells), CD3 (T cells) and B220 (B lymphocytes). The obtained cells were counted, tested for viability and suspended in PBS for injection. Intraocular injection of 200X105 WBMC or Fr25lin- to neonate mice aged 4-12 days was performed under isoflurane anesthesia.
Donors cells that constitutively express GFP were detected in the neonate eyes 1 month following transplantation of either WBMC or Fr25lin- cells. The cells that incorporated into the developing retinae were mainly located in the RGC layer and showed green cytoplasm. Differentiation studies demonstrated neuronal and glial, and only few inflammatory markers.
Neonates survived intraocular injection of putative stem cells from GFP+ adult bone marrow donors. The eyes maintained their normal anatomy. A few cells, which were incorporated into the retina, as detected 1 month following transplantation, demonstrated neuronal and glial differentiation. The survival and differentiation of the GFP + cells for longer than 6 months is currently under investigation.
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